Process for synthesizing active esters of carboxylic acids

ABSTRACT

The invention relates to a new process for preparing active esters or carboxylic acids, which consists in reacting a carboxylic acid, in the presence of an agent for binding hydrohalic acid, with a carbonate of formula: ##STR1## in which R 1  denotes either a radical of formula ##STR2## in which R 3  and R 4 , which may be identical or different, are not hydrogen atoms and denote organic radicals which may be substituted or unsubstituted and saturated or unsaturated, and may be or may not be bound to a polymer, and which can be joined together to form a hetero-cyclic system with the nitrogen atom, 
     or a substituted or unsubstituted aryl radical which may or may not be bound to a polymer, 
     R 2  denotes a hydrogen atom, an aliphatic or cycloaliphatic radical which may be substituted or unsubstituted and saturated or unsaturated, or a substituted or unsubstituted aromatic radical, 
     and X denotes a halogen atom. 
     This process is especially useful for the synthesis of active esters of N-protected amino acids. The invention also relates to the new carbonates described above and the method of producing them, which consists in reacting an alpha-halogenated chloroformate of formula: 
     
         R.sup.2 --CHX--O--COCl 
    
     with an alcohol of formula R 1  OH in an inert solvent medium of the presence of an organic or inorganic base.

The invention relates to a new synthesis of active esters of carboxylicacids. It also relates to the new alpha-halogenated carbonates which areuseful for this synthesis, and to the method of producing them.

Active esters of carboxylic acids are much sought after, especially inthe field of peptide synthesis. Thus, esterified, N-protected aminoacids are stable products which may be stored and marketed. It issubsequently only necessary to react them with an amino compound inorder to form the peptide bond.

The various types of active esters are well described in the literature(Houben Weyl Vol. 15, 2nd part, or Specialist Periodical Reports"Amino-acides Peptides and Protein" Vol. 1-14).

Among the most common esters, substituted aryl esters, such as2,4,5-trichlorophenyl esters, and esters of hydroxyamines, such asN-succinimidyl esters, may be mentioned.

These active esters can also be used for coupling biologically activemolecules to proteins or to solid or liquid supports (German PatentApplication No. 2,631,656, European Patent Application No. 2,677).

They are also used for grafting amino acids onto biologically activemolecules (European Patent Application No. 10,297).

A number of processes have already been proposed for preparing them[Tetrahedron 36 2409 (1980)].

The most widely used process consists of condensing the acid and thehydroxyl compound in the presence of dicyclohexylcarbodiimide as adehydrating agent, but the yields are variable and there is formed, asby-product, dicyclohexylurea, which is very difficult to removecompletely, and N-acylureas [Chem. Ber. 100, 16 (1967)].

According to another process, the hydroxylated compound is reacted withthe mixed anhydride formed from the acid to be esterified and acarboxylic, carbonic or phosphoric acid chloride. Two stages are thengenerally necessary. Acid chlorides are unpleasant reagents to handleand are frequently toxic.

It is also possible to obtain active esters by reaction of the acid witha symmetrical carbonate formed from the hydroxylated compound. However,one equivalent of the hydroxylated compound is then lost, and this isfrequently an expensive product which it is difficult to remove from themedium and recover.

It has also been proposed to replace the hydroxylated compound, in thecase of 2,4-dinitrophenol, by its fluoride, which reacts with the sodiumsalt of the acid. However, this method is highly specific.

Some authors have, in fact, employed more sophisticated reagents such as2-halopyridinium salts or the triphenylphosphine/diethylazodicarboxylate system.

The very large number of processes proposed in the literature forproducing these active esters tends to substantiate the view that noneis entirely satisfactory, and that, in view of the value of thesecompounds, a better solution is still being sought.

The object of the invention is specifically to provide a process inwhich this expectation is fulfilled.

According to the process of the invention, a carboxylic acid is reacted,in the presence of an agent for binding hydrohalic acid, with analpha-halogenated carbonate of the formula ##STR3## in which R¹ denoteseither a radical of formula ##STR4## in which R³ and R⁴, which may beidentical or different, are not hydrogen atoms and denote highly diverseorganic radicals which may be substituted or unsubstituted and saturatedor unsaturated, and may or may not be bound to a polymer, and which canbe joined together to form a heterocyclic system with the nitrogen atom,

or a substituted or unsubstituted aryl radical which may or may not bebound to a polymer,

R² denotes a hydrogen atom, an aliphatic or cycloaliphatic radical whichmay be substituted or unsubstituted and saturated or unsaturated, or asubstituted or unsubstituted aromatic radical,

and X denotes a halogen atom.

The reaction scheme is as follows: ##STR5## This reaction is surprisingsince, in effect, it is known that alpha-halogenated carbonates reactwith acids to give ester carbonates according to the scheme: ##STR6##This reaction is described, in particular, in Patents FR No. 2,201,870and 82,404, and is very useful for improving the efficacy of certaindrugs.

The mechanism of the reaction according to the invention is completelydifferent, and an advantage of the invention is that the conditions havebeen discovered whereby active esters may be obtained.

The process according to the invention is applied to a very large numberof simple or complex acids, saturated or unsaturated aliphatic,cycloaliphatic and heterocyclic acids, as well as aromatic acids. By wayof example, acetic, acrylic, thiophenecarboxylic and benzoic acids maybe mentioned.

It is especially useful for the esterification of N-protected natural orsynthetic amino acids.

If the acids bear especially reactive groups, it is preferable toprotect these but it is generally not necessary.

The alpha-halogenated carbonates of formula ##STR7## which R¹ and R²have the above meanings and which are reacted with the acids, are newcompounds which also form the subject of the present invention.

R³ and R⁴ can, for example, be aliphatic or aromatic radicals, and theyare frequently joined together and R¹ then denotes one of a number ofheterocyclic radicals. This heterocyclic system can comprise one or morehetero atoms which may be identical or different, and it can besaturated or unsaturated, may or may not be condensed with an aromaticsystem and can be substituted or unsubstituted.

R¹ also frequently denotes a substituted or unsubstituted phenylradical.

The substituents of R¹ can be hydrocarbon groups, halogen atoms,especially chlorine or fluorine atoms, or functional groups such asnitro.

The radicals which are regarded as good activating groups in the activeesters are the preferred radicals. These are, for example,N-succinimidyl, para-nitrophenyl, 2,4-dinitrophenyl,2,4,5-trichlorophenyl, pentachloro or -fluorophenyl, and N-phthalimidylradicals.

X is a halogen atom, a fluorine, chlorine, bromine or iodine atom, andis preferably chlorine.

R² generally denotes a C₁ to C₅ aliphatic radical, especially a methylradical which can be substituted with one or more halogen atoms, or aphenyl radical which is unsubstituted or substituted with, for example,one or more halogen atoms.

The trichloromethyl radical is very suitable.

The alpha-halogenated carbonates of the invention are generallycrystalline, stable at room temperature and of little or no toxicity.

The method of producing them, which also forms the subject of theinvention, will be described shortly.

One of the applications thereof is the preparation of active esters asdescribed above.

In this application, one equivalent of carbonate is generally used forone equivalent of acid. If necessary, it is possible to depart from thisproportion.

The carbonate and the acid are reacted in a solvent which is inert withrespect to them. This solvent is generally chosen from cyclic or acyclicethers, such as tetrahydrofuran or dioxane, esters such as, for example,ethyl acetate, nitriles such as acetonitrile, alcohols such as, forexample, isopropanol, amides such as, for example, dimethylformamide andketones such as, for example, acetone. Tetrahydrofuran is very suitable.

The presence of an agent for binding hydrohalic acid is necessary, toremove the acid which forms during the reaction. All the customary meanscan be employed. This will generally be an inorganic base, for examplepotassium carbonate, or an organic base, for example a tertiary amine.

Among preferred bases, N-methylmorpholine or triethylamine may bementioned.

One equivalent of base is generally used per equivalent of acid, but anexcess of base is no disadvantage.

The reaction temperature is generally between -20° and 100° C. It ischosen according to the reactivity of the carbonate used. It is mostfrequently between 20° and 30° C.

As regards the reaction time, this varies from a few minutes to a fewhours. In general, two hours are sufficient to obtain good results. Asimple means for establishing the completion of the reaction is toobserve the cessation in the evolution of carbon dioxide.

The esterified acid obtained is purified by conventional, simplemethods. The by-products of the reaction, which are the hydrochloride ofthe base used and the aldehyde, are readily removed, for example bywashing with water. Then, by evaporation of the solvent, a product isobtained which is already of high purity and which can be recrystallisedin a suitable solvent. The yields are excellent.

The invention also relates to the method of producing the newalpha-halogenated carbonates described above.

This method of production consists in reacting the hydroxylated compoundof formula R¹ OH, in which R¹ has the meaning given above, with analpha-halogenated chloroformate of formula ##STR8## in which R² and Xhave the meaning given above, in an inert solvent medium at atemperature between -20° and 100° C., in the presence of an organic orinorganic base.

The reaction is as follows: ##STR9##

The starting hydroxylated compounds and alpha-halogenated compounds aregenerally commercially available or are manufactured according to knownmethods. For example, the alpha-halogenated chloroformates are preparedby halogenation of the corresponding chloroformates, or as described inFrench Patent Applications Nos. 2,482,587 and 2,532,933, and EuropeanPatent Application No. 108,547.

As an organic or inorganic base, a base is generally employed which isknown to bind hydrochloric acid. Pyridine or triethylamine is preferablyused.

It is preferable to add the base to the mixture of the reagents, andalso to add it gradually.

A stoichiometric amount of the base and the reactive components isgenerally used, but an excess of alcohol can be employed.

The solvent medium consists of one or more solvents which are inerttowards the reagents. Chlorinated aliphatic solvents, for exampledichloromethane and dichloroethane, cyclic or acyclic ethers, ketonessuch as acetone or 2-butanone, nitriles, esters and aliphatic oraromatic hydrocarbons are preferably chosen.

When certain hydroxylated compounds such as phenols are reacted, it ispreferable to choose a solvent system in which the phenol is onlyslightly soluble and the hydrochloride of the base is insoluble. Abenzene/petroleum ether mixture, for example, is very suitable.

The reaction temperature is preferably between -10° and 40° C.

When the introduction of the reagents is complete, the mixture isgenerally brought to room temperature. The reaction time is frequentlybetween a few minutes and a few hours.

The carbonate obtained is readily isolated by the usual methods. Forexample, the mixture is washed wih iced water or acid sodium sulphate,or filtered on a celite bed and the solvent or solvents then evaporated.The crystals generally obtained can be recrystallised in theconventional manner.

Thus, the invention enables new carbonates to be prepared in a simplemanner without a special installation. Due to the originality of theirstructure, these new carbonates may form the subject of highly diverseapplications.

The application to the esterification of acids, which forms the subjectof the present invention, is a very useful application. The activeesters of the acids are thereby obtained without great difficulty andwithout racemization, by using in a simple manner reagents which areeasy to obtain and handle. The by-products of the reaction, thehydrochloride of the base used and aldehyde, are readily removed. Theyields are very good and the products of high purity.

The examples which follow are given by way of illustration.

EXAMPLES 1 TO 10: SYNTHESIS OF ALPHA-HALOGENATED CARBONATES

The carbonates are prepared according to one or more of the followingmethods. The results are collated in Table 1.

METHOD A

12.35 g (0.05 mol) of 1,2,2,2-tetrachloroethyl chloroformate are addedin a single portion to a solution of 5.75 g (0.05 mol) ofN-hydroxysuccinimide in dichloromethane (50 ml). The mixture is cooledto 0° C. and 4 g (0.05 mol) of pyridine is added dropwise. When theaddition is complete, the mixture is allowed to return to roomtemperature and stirred for 3 hours. 20 ml of iced water are then added,and the organic phase is separated and washed with 20-ml portions oficed water as many times as are necessary until the pH is no longer acid(generally 3 to 4 times). The organic phase is dried over magnesiumsulphate and evaporated, and a white solid is obtained which iscrystallised in petroleum ether. 13.5 g (yield Y=83%) of the expectedcarbonate are collected. This recrystallisation can be obmitted; theyield is then 94% and the product is of satisfactory purity.

METHOD B

The procedure is as in Method A, replacing pyridine by triethylamine.The reaction is carried out for one hour at 0° C. and then one hour atroom temperature. In this case, the organic phase is washed rapidly withsaturated NaHSO₄ solution. After evaporation, the N-hydroxysuccinimidecarbonate is crystallised in ethyl ether (Y=83%).

METHOD C

27 g (0.11 mol) of 1,2,2,2-tetrachloroethylchloroformate are added to asuspension of 18.4 g (0.1 mol) of 2,4-dinitrophenol in 150 ml of benzeneand 150 ml of petroleum ether. The mixture is cooled to 0° C. and 11 g(0.11 mol) of triethylamine are added dropwise while vigorous stirringis maintained. The mixture is stirred for 4 hours at 20° C. and thenfiltered on a celite bed. The solvents are evaporated and 26 g (Y=66%)of white crystals are obtained after being washed with a littlepetroleum ether.

    TABLE 1          Crystallisation M.p. or B.p.    Ex Product Method Yield solvent mm     Hg °C. .sup.1      H NMR ppm IR cm.sup.-1 Elementary analysis                     1      ##STR10##      AB 8383 Petroleum etherEthyl ether 104108 2,9(s)6,6(s) 1760 C 26,00; H     1,74; N 4,10; O 23,54; Cl 43,55;(calculated: C 25,87; H 1,55; N 4,31;O     24,6; Cl 43,55;      2     ##STR11##      A 64 Petroleum ether 71 6,73(s)7,4(s) 1800 C 26,61; H 1,02; O 11,56; Cl     60,80;(calculated C 26,54; H 0,74; O 11,78; Cl 60,93)      3     ##STR12##      A 92% crude70% dist. -- B.p. 0,02 =150-155 6,7(s)7,4(s)7,56(s) 1795 C     26,67; H 0,90; O 11,95; Cl 60,64;(calculated: C 26,54; H 0,74;O 11,78;     Cl 60,93;      4     ##STR13##      A 98 AcOEt 120 6,7(s) 1800 C 22,40; H trace; Cl 67,15;(calculated: C     22,66; H 0,27; Cl 67,05      5     ##STR14##      A 91 -- B.p. 0,05 = 80 6,7(s) 1800 C 27,60; H 0,47; Cl 35,80; F     24,56;(calculated C 27,44; H 0,26; Cl 36,00; F 24,12);      6     ##STR15##      C 75 Petroleum ether B.p. 0,05 = 165M.p. 69 6,7(s)7,5(d)8,36(d) 1790 C     30,87; H 1,53; N 3,81;O 22,23; Cl 41,15; (calculated C 30,97; H 1,44; N     4,01; O 22,92; Cl 40,64)      7     ##STR16##      C 66 Petroleum ether 121-122 6,73(s)8,60(s)9,06(d)7,66(d) 1795 C 27,49;     H 1,04; N 6,98;O 27,99; Cl 35,95;(calculated C 27,44; H 1,02;N 7; O     28,43; Cl 36,00)       8     ##STR17##      A 75  B.p. 0,05 =115-131 2,8(s)7,2(s)7,3 to8(m) 1750 M.sup.+  + 18 =     335      9     ##STR18##      B 85 CH.sub.2 Cl.sub.2 145-147 7,04(s)7,6(d) 7,8(dd)8,0(d)8,2(d)     17701790 C 31,38; H 1,37; N 12,16;  Cl 41,03;(calculated C 31,30; H     1,45; N 12,17;Cl 41,16)  10      ##STR19##      A 86 Petroleum ether 105-106 6,4(q)1,9(d)2,8(s) 1750 C 37,80; H 3,64; N     6,24;O 36,68; Cl 15,88;(calculated C 37,94; H 3,64; N 6,32;O 36,10; Cl     16,00

EXAMPLE 11: PREPARATION OF N-SUCCINIMIDYLN-TERT-BUTYLOXYCARBONYL-L-ALANINATE

0.95 g (5 mmol) of N-tert-butyloxycarbonyl-L-alanine and 0.66 ml (5.5mmol) of N-methylmorpholine are dissolved in 6 ml of THF, and 1.8 g (5.5mmol) of N-succinimidyl 1,2,2,2-tetrachloroethyl carbonate is added in asingle portion. The mixture is stirred for two hours at 20° C.Approximately 25 ml of ethyl acetate are added and the organic phase iswashed rapidly with N HCl solution, then with potassium bicarbonatesolution and finally twice with water. The organic phase is dried overmagnesium sulphate and evaporated. The residue is crystallised in anethyl acetate/petroleum ether mixture and 1.25 g (Y=87%) of whitecrystals are obtained. Evaporation of the mother liquors andcrystallisation enables a further 0.1 g to be recovered, and this bringsthe yield to 94%.

M.p. (melting point)=158° C. [α]_(D) ²⁰ =50.7 (c=2, dioxane)

EXAMPLE 12: PREPARATION OF N-SUCCINIMIDYL N.sup.α-TERT-BUTYLOXYCARBONYL-Nε-BENZYLOXYCARBONYL-L-LYSINATE

1.64 g (5 mmol) of N-succinimidyl 1,2,2,2-tetrachloroethyl carbonate isadded to a solution of 1.9 g (5 mmol) of N.sup.α -BOC-Nε-Z-lysine and0.7 ml (5 mmol) of triethylamine in 15 ml of THF. The mixture is stirredat room temperature for 2 hours. 20 ml of ethyl acetate are then addedand the organic phase is washed with 0.5N citric acid solution, thenthree times with 5% strength sodium bicarbonate solution and finallywith saturated sodium chloride solution. The organic phase is dried oversodium sulphate and evaporated to dryness. 2.03 g (Y=85%) of whitecrystals are obtained.

M.p 110° C. [α]_(D) ²⁰ =-21.51 (c=2, dioxane)

Analysis calculated for C₂₃ H₃₄ N₃ O₆.5H₂ O: C=56.79; H=6.79; N=8.64Found C=56.32; H=6.44; N=8.77.

EXAMPLES 13 TO 29

Other N-succinimidyl esters are prepared as in Example 11 or 12. All theresults are summarised in Table 2.

                                      TABLE 2                                     __________________________________________________________________________    N--SUCCINIMIDYL ESTERS                                                                                      Litera-                                                                       ture                                                         Yield                                                                             M.p.         yield                                                                             M.p.                                        EX Amino acid                                                                              %   °C.                                                                       [α].sub.D.sup.20 (, solvent)                                                      %   °C.                                                                         [α].sub.D.sup.20 (c,                                                    solvent)      REF                      __________________________________________________________________________    11 BOC-- Ala 94  158                                                                              -50,7 (2, dioxane)                                                                      71  161  -49,0 (2, dioxane)                                                                          1                        12 BOC-- (Z)-- Lys                                                                         85  110                                                                              -21,51 (2, dioxane)                                                                     --  110  -21,12 (2, dioxane)                                                                         2                        13 BOC-- Phe 91  147                                                                              -19,8 (2, dioxane)                                                                      81  152  -20,7 (2,2 dioxane)                                                                         1                        14 BOC-- Gly 91  162                                                                              --        62  168  --            1                        15 BOC-- Val 92  125                                                                              -37,4 (2, dioxane)                                                                      74  125  -37 (2, dioxane)                                                                            1                        16 BOC-- Pro 82  133                                                                              -54,7 (2, dioxane)                                                                      74  135  -55,3 (2, dioxane)                                                                          1                        17 BOC-- Tyr 69-74                                                                             181                                                                              -12,12 (1, dioxane)                                                                         180  -12,5 (1, dioxane)                                                                          2                        18 BOC-- Trp 89  140                                                                              -23,13 (2, dioxane)                                                                     37  153  -22,4 (2, dioxane)                                                                          1                        19 BOC-- Met 92  124                                                                              -21,6 (2, dioxane)                                                                      59  128  -20,6 (2, dioxane)                                                                          1                        20 BOC-- (SBzl) Cys                                                                        61  106                                                                              -54,0 (2, dioxane)                                                                      49  117  -54,0 (2, dioxane)                                                                          1                        21 BOC-- (OBzl)-- Ser                                                                      78  -- +5,8 (0,5, dioxane)                                                                     52  112-113                                                                            -[α].sub.D.sup.26 +6,5 (0,5                                             dioxane)      2                        22 BOC-- (OBzl)-- Thr                                                                      85  -- +9,38 (2, dioxane)                                                                      --   94  -8,5 (1, methanol)                                                                          2                        23 BOC-- (OBzl) Tyr                                                                        73  144                                                                              -6,2 (2, dioxane)                                                                       --  149  -6,3 (2, dioxane)                                                                           2                        24 Z-- Ala   86  123                                                                              -36,9 (2, dioxane)                                                                      65  123  -37,2 (2, dioxane)                                                                          1                        25 Z-- Pro   97   90                                                                              -54,9 (2, dioxane)                                                                      74   90  -54 (2, dioxane)                                                                            1                        26 Z-- Met   72  102                                                                              -16,2 (2, dioxane)                                                                      59  102  -15,9 (2, dioxane)                                                                          1                        27 FMOC-- Ala                                                                              70  102                                                                              --        --  --   --            --                       28 FMOC-- Phe                                                                              97  137                                                                              --        --  --   --            --                       29 FMOC-- Pro                                                                              100    --        --  --   --            --                       __________________________________________________________________________     Ref.                                                                          1 J. Am. Chem. Soc. 86, 1839 (1964)                                           2 HoubenWeyl: Methoden der Org. Chemie Vol. 15(2) p. 165 and loc. cit.   

EXAMPLE 30: PREPARATION OF N-SUCCINIMIDYLN-TERT-BUTYLOXYCARBONYL-N-PHENYLALANINATE

To a solution of 1.33 g (5 mmol) of BOC-L-phenylalanine in 6 ml of THF,0.8 g of anhydrous potassium carbonate and 1.8 g of1,2,2,2-tetrachloroethyl N-succinimidyl carbonate are added. The mixtureis stirred for two hours at 20° C. and treated as in Example 11. 1.15 g(Y=63%) of 80C-L-PheOSu is obtained. M.p. 135° C.

If 1 ml of aqueous potassium carbonate solution is used in place ofanhydrous potassium carbonate, 1.3 g (Y=72%) of the above ester isobtained.

EXAMPLE 31: PREPARATION OF N-SUCCINIMIDYLN-TERT-BUTYLOXYCARBONYLGLYCINATE

The procedure is as in Example 11, replacing THF by acetonitrile.Starting with 1.76 g of BOC-glycine, 2.1 g (Y=77%) of BOC-Gly-OSu isobtained.

M.p. 158° C. M.p. lit 168° C.

EXAMPLE 32: PREPARATION OF N-SUCCINIMIDYLN-TERT-BUTYLOXYCARBONYL-N-PHENYLALANINATE

To a solution of 1.33 g (5 mmol) of BOC-L-phenylalanine in 6 ml of THF,0.66 ml of N-methylmorpholine and 1.11 g (5 mmol) of 1-chloroethylN-succinimidyl, carbonate are added. The mixture is stirred for 16 hoursat 20° C. and treated as in Example 11. 1.40 g (Y=77%) of BOC-L-Phe-OSuis obtained.

M.p. 143° C. [α]_(D) ²⁰ =-16.6 (c=2,2 dioxane).

EXAMPLE 33: PREPARATION OF (2-CHLOROPHENYL)CHLOROMETHYLN-TERT-BUTYLOXYCARBONYL-L-ALANINATE

To a solution of 0.95 g (5 mmol) of BOC-L-alanine and 0.66 ml ofN-methylmorpholine in 6 ml of THF, 2.0 g of (2-chlorophenyl)chloromethylN-succinimidyl carbonate are added. The mixture is stirred for 1 hour 30minutes at 20° C. and treated as in Example 11. 1.25 g (Y=90%) ofBOC-L-Ala-OSu is obtained.

M.P. 158°-159° C. [α]_(D) ²⁰ =-49.5 (c=2, dioxane).

EXAMPLES 34 TO 42: PREPARATION OF 2,4,5-TRICHLOROPHENYL ESTERS

These esters are prepared as in Example 11 or 12, starting with1,2,2,2-tetrachloroethyl 2,4,5-trichlorophenyl carbonate. The resultsare collated in Table 3.

EXAMPLE 43: PREPARATION OF PENTACHLOROPHENYLN-TERT-BUTYLOXYCARBONYL-L-ALANINATE

The procedure is as in Example 12. Starting with 0.84 g (5 mmol) ofBOC-L-alanine and 2.27 g (5 mmol) of pentachlorophenyl1,2,2,2-tetrachloroethyl carbonate, 2.06 g (Y=94%) of white crystals areobtained (recrystallisation solvent: ethyl acetate/hexane).

M.p. 170° C. [α]_(D) ²⁰ =-24.47 (c=1, CHCl₃)

literature

M.P. 166° C. [α]_(D) ²⁰ =-22.2 (c=5.1, CHCl₃).

EXAMPLE 44: PREPARATION OF PENTACHLOROPHENYL N.sup.α-TERT-BUTYLOXYCARBONYL-Nε-BENZYLOXYCARBONYL-L-LYSINATE

The procedure is as in Example 12. Starting with 1.9 g of N.sup.α-BOC-Nε-Z-L-lysine, 2.4 g (Y=77%) of N.sup.α -BOC-Nε-Z-L-lysine-OPCP areobtained.

                                      TABLE 3                                     __________________________________________________________________________    2,4,5-TRICHLOROPHENYL ESTERS                                                                                 Litera-                                                     Yield                                                                             M.p.          ture                                                                              M.p.                                                                             [α].sub.D.sup.20 (c, solvent      EX Amino acid                                                                              %   °C.                                                                       [α].sub.D.sup.20 (c, solvent)                                                      yield                                                                             °C.                                                                       literature Ref                          __________________________________________________________________________    34 BOC-- Ala 79  82 -43,5 (2, acetic ac.)                                                                    73  82 -44 (2, acetic ac.)                                                                      1                            35 BOC (SBzl) Cys                                                                          60  --            86  77 --         1                            36 BOC (DNP) Hist                                                                          58  98 +1,87 (2, dioxane)                                                                       --  -- --                                      37 BOC-- (OBzl) Glu                                                                        60  107                                                                              -33,5 (2, methanol)                                                                      60  108                                                                              -33,5 (2, methanol)                                                                      2                            38 BOC-- (Z) Lys                                                                           85  99 -19,13 (2, dioxane)                                                                      86  99 --         1                            39 BOC-- Met 79  90 -38,0 (2, DMF)                                                                           71  91 -38,5 (2, DMF)                                                                           1                            40 BOC-- Ser 22  105                                                                              -42,0 (2, DMF)                                                                            5  105                                                                              -41 (2, DMF)                                                                             3                            41 BOC-- (OBz-l) Ser                                                                       77  oil                                                                              + 0,3 (2, dioxane)                                                                       --  -- --         --                           42 BOC-- Tyr.                                                                              73  168                                                                              -31,4 (3,5, acetic ac.)                                                                  40  158                                                                              -26,4 (3,5 acetic                                                                        2c.)                         __________________________________________________________________________     Ref.                                                                          1 J. Chem. Soc (C) (1967)                                                     2 Tetrahedron Suppl. 8,39 (1966)                                              3 J. Med. Chem. 10, 1047 (1977)                                          

M.p. 142° C. [α]_(D) ²⁰ =-16.7 (c=1, CHCl₃).

literature:

M.p. 141° C. [α]_(D) ²⁰ =-15.0 (c=4.99, CHCl₃).

EXAMPLE 45: PREPARATION OF 4-NITROPHENYLN-TERT-BUTYLOXYCARBONYL-L-PHENYLALANINATE

0.73 g (2.75 mmol) of BOC-L-phenylalanine and 0.3 ml ofN-methylmorpholine are dissolved in 6 ml of THF, and 87 g (2.5 mmol) of1,2,2,2-tetrachloroethyl 4-nitrophenyl carbonate are added. The mixtureis stirred for one hour at room temperature 15 ml of ethyl acetate arethen added and the mixture is washed with normal hydrochloric acidsolution and then with saturated sodium chloride solution. The organicphase is dried over magnesium sulphate. It is evaporated to dryness andthe product is crystallised in 95% strength ethanol. 0.6 g (Y=62%) ofwhite crystals are obtained.

M.p. 118° C. [α]_(D) ²⁰ =-20.9 (c=2, DMF).

literature:

M.p. 132° C. [α]_(D) ²⁰ =-21 (c=2, DMF).

EXAMPLE 46: PREPARATION OF 2,4-DINITROPHENYLN-TERT-BUTYLOXYCARBONYL-L-ALANINATE

The procedure is as in the example above. Starting with 0.945 g (5 mmol)of N-BOC-L-alanine, 1.45 g (Y=82%) of N-BOC-L-Ala-O-2,4-DNP,crystallised in 95% strength ethyl alcohol, is obtained.

M.p. 95°-96° C. [α]_(D) ²⁰ =-51.5 (c=0.2, DMF).

EXAMPLES 47 TO 54: PREPARATIONS OF PENTAFLUOROPHENYL ESTERS

The procedure is as in Example 11. The results are collated in Table 4.

                                      TABLE 4                                     __________________________________________________________________________    PENTAFLUOROPHENYL ESTERS                                                                      Lit.                                                                   Yield                                                                             M.p.                                                                             M.p.         Literature                                       EX Amino acids                                                                         %   °C.                                                                       °C.                                                                       [α].sub.D.sup.20 (c, solvent)                                                     [α].sub.D.sup.20 (c,                                                              REF.ent)                               __________________________________________________________________________    47 BOC-- Ala                                                                           90  85 83 -34,6 (1, dioxane)                                                                      -31,2 (1, dioxane)                                                                      1                                      48 BOC-- Gly                                                                           90  80 80 --        --                                               49 BOC-- Phe                                                                           86  112                                                                              112                                                                              -16,9 (1, dioxane)                                                                      -26,9 (1, dioxane)                                                                      1                                      50 BOC-- Val                                                                           94  64 64 -31,8 (1, dioxane)                                                                      -18,4 (1, dioxane)                                                                      1                                      51 BOC-- Pro                                                                           98  50 51 -48,2 (1, dioxane)                                                                      -53 (1, dioxane)                                                                        1                                      52 BOC-- Trp                                                                           91  111                                                                              111                                                                              -12,8 (1, dioxane)                                                                      -28,1 (1, dioxane)                                                                      1                                      53 BOC-- Met                                                                           81  79 80 -20,7 (1, dioxane)                                                                      -22,5 (1, dioxane)                                                                      1                                      54 Z-- Pro                                                                             100 oil                                                                              oil                                                                              -55,08 (1, dioxane)                                                                     -55,05 (1, dioxane)                                                                     1                                      __________________________________________________________________________     Ref.                                                                          1 Ann. 1421 (1973)                                                       

EXAMPLE 55: PREPARATION OF PENTAFLUOROPHENYLN-TERT-BUTYLOXYCARBONYL-L-PHENYLALANINATE

The procedure is as in Example 48, but varying the reaction solvent. Theresults obtained are collated in the following table:

    ______________________________________                                        SOLVENT        YIELD      M.P. °C.                                                                        [α].sup.20                           ______________________________________                                        THF            86         112      -16.9                                      DIOXANE        86         113      -16.4                                      ETHYL ACETATE  83         112      -15.4                                      DMF            65         107      -14.2                                      ______________________________________                                    

EXAMPLES 56 AND 57

These examples illustrate the use of the active esters of the aminoacids in the synthesis of di- and tripeptides.

56. Preparation of ethyl N-BOC-L-phenylalanylglycinate (BOC-Phe-Gly-OEt)

To a solution of 0.86 g (2 mmol) of pentafluorophenylN-t-butyloxycarbonyl-L-phenylalaninate, prepared according to Example48, in 10 ml of dioxane, 0.28 g (2.2 mmol) of ethyl glycinate(hydrochloride) and 0.31 g (3 mmol) of triethylamine are addedsimultaneously. The mixture is stirred for one hour at room temperature.It is diluted with 10 ml of ethyl acetate and washed with aqueous 0.1Nhydrochloric acid, potassium bicarbonate and sodium chloride solutions.The organic phase is dried over magnesium sulphate and evaporated. Theresidue is taken up with a 4:6 ethyl acetate/hexane mixture and filteredon silica. After evaporation, 0.53 g (Y=76%) of BOC-Phe-Gly-OEt isobtained.

M.p. 89° C. [α]_(D) ²⁰ =-4.8 (c=5.0, EtOH)

M.p._(lit) =89.5° C. [α]_(D) ²⁰ -4.2 (c=5.0, EtOH) [J. Amer. Chem. Soc.82, 4596 (1960)].

57 Preparation of ethyl N-benzyloxycarbonylprolinylglycylglycinate

    Z-Pro-GlyOH→Z-Pro-Gly-OPFP→Z-Pro-Gly-Gly-OEt

1.4 g (3.6 mmol) of 1,2,2,2-tetrachloroethyl pentafluorophenyl carbonateis added to a solution of 1.07 g (3.6 mmol) ofN-benzyloxycarbonylprolinylglycine and 0.4 ml of N-methylmorpholine in 5ml of dioxane. The mixture is stirred for one hour at 20° C. and treatedas in Example 55. 0.82 g (Y=45%) of Z-Pro-Gly-OPFP is obtained:

[α]_(D) ²⁰ =-36.8 (c=1, dioxane).

This 0.8 g is dissolved in 10 ml of dioxane, and 0.23 g (1.8 mmol) ofethyl glycinate hydrochloride and 0.25 g of triethylamine are addedsimultaneously. The procedure is then as in Example 55. 0.55 g (Y=94%)of Z-Pro-Gly-Gly-OEt is obtained.

M.p. 95°-96° C. [α]_(D) ²⁰ =-21.4 (c=1, EtOH)

which is recrystallised in an ethyl acetate/hexane mixture.

M.p. 117°-118° C. [α]_(D) ²⁰ =-24.6 (c=1, EtOH)

M.p._(lit) 120° C. [α]_(D) ²⁰ =-26 (c=1, EtOH).

EXAMPLE 58: PREPARATION OF N-SUCCINIMIDYL 2,2-DIMETHYLACRYLATE

0.5 g of 2,2-dimethylacrylic acid in 15 ml of THF, 1.63 of1,2,2,2-tetrachloroethyl N-succinimidyl carbonate and 0.7 ml oftriethylamine are added. The mixture is stirred for 2 hours at 20° C.and washed with sodium chloride solution. The organic phase is driedover magnesium sulphate and the solvent is evaporated. 0.6 g (Y=56%) ofthe expected ester is obtained. ¹ H NMR (CDCl₃, TMS): 2.05 (s, CH₃); 2.2(s, CH₃); 2.8 (s, CH₂ CH₂); 5.9 (m, H--C═).

EXAMPLE 59: PREPARATION OF N-SUCCINIMIDYL 2-THIOPHENECARBOXYLATE

The procedure is as in the example above. Starting with 0.64 g of2-thiophenecarboxylic acid, 0.62 g (Y=51%) of the expected ester isobtained.

M.p 140°-142° C.

¹ H NMR (CDCl₃, TMS): 2.8 (CH₂ CH₂), 7.0-7.2 (m, H--C═); 7.6 (m, H--C═);7.8 (m, H--C═).

We claim:
 1. A method for the preparation of an active ester of formula##STR20## from an N-blocked aminocacid of formula YCOOH, wherein Ycontains an N-protected amino group, which consists of reacting saidaminoacid with a carbonate of formula ##STR21## wherein R¹ is:(1) anN-succinimidyl (2) an N-phthalimidyl (3) 1-benzotriazolyl radical; (4)phenyl substituted by 1-5 chlorine, fluorine or one or two nitro groups;R² is: a saturated alkyl of 1 to 5 carbon atoms, unsubstituted orsubstituted by chlorine atoms, or an unsubstituted phenyl or phenylsubstituted by chlorine atoms,in the presence of an agent for bindinghydrochloric acid which is a tertiary amine or an alkali metal carbonatein the presence of an inert solvent which is a member selected from thegroup consisting of cyclic or acyclic ethers, esters, nitriles,alcohols, amides and ketones wherein the reaction proceeds as follows:##STR22## and isolating said active ester from the reaction mixture. 2.The process according to claim 1, wherein the carboxylic acid is anamino acid, the amino group of said amino acid being protected. 3.Process according to claim 1, wherein R¹ is a N-succinimidyl,2,4,5-trichlorophenyl, 2,4,6-trichlorophenyl, pentachloro- orpentafluorophenyl, 4-nitrophenyl, 2,4-dinitrophenyl or N-phthalimidylgroup.
 4. The process according to claim 1, wherein the tertiary amineis triethylamine or N-methylmorpholine.
 5. The process according toclaim 1, wherein the solvent is tetrahydrofuran.
 6. The processaccording to claim 1, wherein the radical R² is trichloromethyl.
 7. Theprocess according to claim 1, wherein the reaction temperature isbetween -20° C. and 100° C.
 8. The process according to claim 2, whereinsaid amino acid is alanine, lysine, phenylalanine, glycine, valine,proline, tyrosine, tryptophan, methionine, cysteine, serine, threonine,histidine or glutamic acid and the amino group of said aminoacid isprotected by a benzyloxycarbonyl group, tertiary butyloxycarbonyl,fluorenylmethyloxycarbonyl.
 9. A method for the preparation of an activeester of formula ##STR23## from a carboxylic acid of formula RCOOH,wherein said carboxylic acid is acetic acid, an acrylic acid,thiophenecarboxylic acid or benzoic acid, which consists of reactingsaid carboxylic acid with a carbonate of formula ##STR24## wherein R¹is:(1) an N-succinimidyl; (2) an N-phthalimidyl; (3) benzotriazolylradical; (4) phenyl substituted by 1-5 chlorine, fluorine or one or twonitro groups; R² is: a saturated alkyl of 1 to 5 carbon atoms,unsubstituted or substituted by chlorine atoms, or an unsubstitutedphenyl or phenyl substituted by chlorine atoms,in the presence of anagent for binding hydochloric acid which is a tertiary amine or analkaline metal carbonate in the presence of an inert solvent which is amember selected from the group consisting of cyclic or acyclic ethers,esters, nitriles, alcohols, amides and ketones, wherein said reactionproceeds as follows: ##STR25## and isolating said active ester from thereaction mixture.